PROJECT SUMMARY: Metabolic syndrome affects over 25% of adults in the US and is difficult to treat due to its multifactorial and complex nature. Glucose dysregulation, either excess or deficiency of glucose levels, is a hallmark of metabolic syndrome with both short- and long-term detrimental physiological effects. Hepatic gluconeogenesis, the endogenous production of glucose in the liver, is critical for maintenance of glucose homeostasis during periods of prolonged fasting. Although gluconeogenesis is influenced by a myriad of systems, strong evidence suggests that host circadian rhythms and cues provided by the gut microbiome significantly contribute to this process. Disruption of either system leads to aberrant hepatic gluconeogenesis, although few mechanistic insights provide an explanation for how these consequences arise and how the two systems are connected. Using novel experimental approaches, I will tease apart these relationships to define the specific mechanisms of action of the hepatic core circadian clock and gut microbiota in regulating glucose homeostasis. Thus, I will address the unmet need for targeted and effective therapeutic interventions for metabolic syndrome. My preliminary data show that targeted deletion of the hepatic core circadian clock gene Bmal1 leads to increased glucose clearance and reduced gluconeogenesis in Specific Pathogen Free (SPF) but not germ-free (GF) mice. This phenotypic difference can be restored by transplantation of a full microbial community into GF hepatic Bmal1 deficient mice, implying that gut microbes are necessary and sufficient for proper hepatic Bmal1 regulation of gluconeogenesis. Microbial 16S rRNA sequencing of stool serially collected over 48 hours reveals that mice lacking hepatic Bmal1 exhibit nearly twice the number of oscillating Clostridia taxa, suggesting changes in microbial community dynamics and function could feed back to alter host metabolism. These data led me to hypothesize that hepatic gluconeogenesis is driven by bidirectional interactions between the hepatic circadian clock and diurnal patterns of specific classes of gut microbes and metabolites. I will utilize both SPF and GF Bmal1-floxed Albumin-Cre transgenic mice, where Bmal1 is deleted only in hepatocytes. Two central aims are proposed: 1) Examine the central role of the liver clock in transducing gut microbial cues that regulate host hepatic GNG and glucose metabolism, and 2) Identify how known gut microbial products that have been shown to modulate liver clock function mediate clock-controlled hepatic GNG and systemic glucose regulation. I will apply in vivo experimental models, gnotobiotic technology, functional metagenomic analyses, and murine conventionalization experiments. This strategy will fill crucial gaps in knowledge relevant to the interactions between gut microbes and peripheral circadian clocks, as well as mechanisms governing how each system imposes unique influence on hepatic gluconeogenesis. These studies serve as an outstanding training vehicle for me to develop the experimental and critical thinking skills necessary to become a productive and independent researcher focused on the microbial basis of host metabolism.